Substituted benzopyrans

ABSTRACT

PCT No. PCT/US96/20607 Sec. 371 Date Mar. 11, 1998 Sec. 102(e) Date Mar. 11, 1998 PCT Filed Dec. 20, 1996 PCT Pub. No. WO97/25040 PCT Pub. Date Jul. 17, 1997This invention covers certain hydroxy-substituted benzopyrans which are useful for treating asthma and other diseases involving leukotriene-related disease states.

This application is a 371 of PCT/U.S. Ser. No. 96/20607 filed Dec. 20,1996.

SCOPE OF THE INVENTION

This invention covers certain benzopyran compounds which have activityas leukotriene antagonists and are active metabolites of an anti-asthmadrug named pranlukast.

AREA OF THE INVENTION

Substituted benzopyran compound which have activity as leukotrieneantagonists are known in the art. For example U.S. Pat. No. 4,780,469which corresponds to Japanese patent 1741466 and EP patent EP O 173516-A discloses a class of benzopyrans which are described asantagonists of the leukotrienes, particularly the peptido-leukotrienesLTC₄, LTD₄, and LTE₄. As such, these compounds are useful for treating ahost of diseases associated with modification of the metabolic pathwaywhich has these leukotrienes as intermediates. An example of suchdiseases is asthma.

This invention relates to an benzylhydroxybenzopyran andarylhydroxybenzopyran as illustrated by Formula I and II respectively.##STR1##

Formula I is named 4-oxo-8-4-(4-hydroxy-4-phenylbutoxy)benzoylamino!-2-tetrazol-5-yl-4H-1-benzopyran.

Formula II is named 4-oxo-8-4-(4-hydroxy-4-(4-hydroxyphenyl)butoxy)benzoylarnino!-2-tetrazol-5-yl-4H-1-benzopyran.

This invention also relates to an essentially pure preparation of acompound of formula I or II, or a pharmaceutical composition of oneand/or the other in combination with a pharmaceutically acceptableexcipient. It also relates to a method for manufacturing a medicarnentfor treating a disease such as asthma using fornula I or II, or anessentially pure form thereof. These compounds are useful in treatingasthma. Hydrates, solvates, tautomeric forms, isomers and polymorphs ofthese compounds are also included with the scope of this invention.

DETAILED DESCRIPTION OF THE INVENTION

These two compounds may be prepared by synthetic chemistry means or canbe obtained by extracting them from biological samples collected frommammals which have been given the compound 8-p-(4-phenylbutyloxy)benzoyl!amino-2-tetrazol-5'-yl)-4-oxo-4H-1-benzopyran(pranlukast). A synthetic process for one of the compounds is givenbelow; the other can be isolated from biological sources as furtherdescribed below. The compound named in the preceeding sentence can bemade by the chemistries set out in U.S. Pat. No. 4,780,469 whichcorresponds to Japanese patent 1741466 and EP patent EP O 173 516-A.

The benzylic hydroxylation (Formula (I)) of pranlukast appears to be amajor route of metabolism in humans. This metabolism results inmeasurable plasma concentrations in humans. Results from a humanradiolabel study suggests that 98.4% of the administered radiolabel isrecovered in feces (˜84% as pranlukast, 6.6% as an arylhydroxymetabolite Formula (II)! and about 6% as benzylhydroxy metaboliteFormula (I)!. The arylhydroxy and benzylhydroxy metabolic pathwaysappear to be the two major metabolitic routes of pranlukast metabolismin humans and when considering the poor absorption of pranlukast, thequantities of these metabolites measured in feces may represent thetotal of absorbed and metabolized parent.

In incubations of liver microsomes from rats or mice, the arylhydroxyand benzylhydroxy metabolites are detectable when using highconcentrations of pranlukast (50 uM), but hydrolysis of the amide bondin the center of the molecule is the predominant metabolic pathway inthese rodent incubations. Liver microsomes from dogs form thearylhydroxy species and only very small amounts of the benzyl species.

The arylhydroxy and benzylhydroxy metabolites are measurable in humanplasma, but only limited data are available from the singleradiolabelled study. The arylhydroxy metabolite appears to be sulfatedin humans but there is no evidence for sulfation of the benzylhydroxymetabolite. From an evaluation of a battery of human liver microsomes,it was evident that some human livers have enhanced activity towardsbenzylic hydroxylation. It appears that benzylic and aryl hydroxylationare performed by different P450 enzymes but which enzyme(s) is currentlynot known.

As regards the pharmacological activity of Formula I and II both havebeen found to be active in assays predictive of utility in treatingasthma and other diseases which can be treated by pranlukast.

It will be recognized that both compounds may exist in both racemic andoptically active forms. Both forms are to be considered to be within thescope of the present invention.

Pharmaceutically acceptable salts are prepared in a standard manner. Thehydrogen on the tetrazole ring is sufficiently acidic so as to formsalts at a pH of about 6-7 or higher.

Pharmaceutical compositions of the present invention comprise apharmaceutical carrier or diluent and some amount of a compound of theformula (I) or (II). The compound may be present in an amount to effecta physiological response, or it may be present in a lesser amount suchthat the user will need to take two or more units of the composition toeffect the treatment intended. These compositions may be made up as asolid, liquid or in a gaseous form. Or one of these three forms may betransformed to another at the time of being administered such as when asolid is delivered by aerosol means, or when a liquid is delivered as aspray or aerosol.

The nature of the composition and the pharmaceutical carrier or diluentwill, of course, depend upon the intended route of administration, forexample parenterally, topically, orally or by inhalation.

For topical administration the pharmaceutical composition will be in theform of a cream, ointment, liniment, lotion, pastes, aerosols, and dropssuitable for administration to the skin, eye, ear, or nose.

For parenteral administration the pharmaceutical composition will be inthe form of a sterile injectable liquid such as an ampule or an aqueousor non-aqueous liquid suspension.

For oral administration the pharmaceutical composition will be in theform of a tablet, capsule, powder, pellet, atroche, lozenge, syrup,liquid, or emulsion.

When the pharmaceutical composition is employed in the form of asolution or suspension, examples of appropriate pharmaceutical carriersor diluents include: for aqueous systems, water; for non-aqueoussystems, ethanol, glycerin, propylene glycol, corn oil, cottonseed oil,peanut oil, sesame oil, liquid parafins and mixtures thereof with water;for solid systems, lactose, kaolin and mannitol; and for aerosolsystems, dichlorodifluoromethane, chlorotrifluoroethane and compressedcarbon dioxide. Also, in addition to the pharmaceutical carrier ordiluent, the instant compositions may include other ingredients such asstabilizers, antioxidants, preservatives, lubricants, suspending agents,viscosity modifiers and the like, provided that the additionalingredients do not have a detrimental effect on the therapeutic actionof the instant compositions.

The pharmaceutical preparations thus described are made following theconventional techniques of the pharmaceutical chemist as appropriate tothe desired end product.

In these compositions, the amount of carrier or diluent will vary butpreferably will be the major proportion of a suspension or solution ofthe active ingredient. When the diluent is a solid it may be present inlesser, equal or greater amounts than the solid active ingredient.

Usually a compound of formula (I) or (II) is administered to a subjectin a composition comprising a nontoxic amount sufficient to produce aninhibition of the symptoms of a disease in which leukotrienes are afactor. Topical formulations will contain between about 0.01 to 5.0% byweight of the active ingredient and will be applied as required as apreventative or curative agent to the affected area. When employed as anoral or other ingested or injected regimen, the dosage of thecomposition is selected from the range of from 50 mg to 1000 mg ofactive ingredient for each administration. For convenience, equal doseswill be administered 1 to 5 times daily with the daily dosage regimenbeing selected from about 50 mg to about 5000 mg.

No unacceptable toxicological effects are expected when these compoundsare administered in accordance with the present invention.

The following examples are given to further illustrate the describedinvention. These examples are intented solely for illustrating theinvention and should not be read to limit the invention in any manner.Reference is made to the claims for what is reserved to the inventorshereunder.

EXAMPLES Characterization of Novel Human Metabolites of PranlukastMethods Preparation, Isolation and Characterisation of Human Metabolites

Preparation of Formula (I) Using Human Hepatic Microsomes:

The incubations were performed with a final volume of 100 mL in ashaking water bath at a temperature of approximately 37° C. Eachincubation contained approximately 400 ug/mL of microsomal protein(H51), 0.5 mL 2 mM pranlukast in a 16% w/v encapsin solution containing1.5% w/v sodium bicarbonate. The incubation volume was adjusted to 75 mLwith 50 mM potassium phosphate buffer (pH 7.4). Following a 5 minutepre-incubation at approximately 37° C., the reaction was initiated bythe addition of 25 mL of pre-warmed cofactor solution (approximately 1.7mg NADP, 7.8 mg glucose 6-phosphate and 6 units of glucose 6-phosphatedehydrogenase per mL of 2% (w/v) sodium hydrogen carbonate). Thereaction was terminated after 30 minutes by adding 100 mL ofacetonitrile and vortex mixed. The precipitate was pelleted bycentrifugation. The putative benzyl hydroxy metabolite was subsequentlyisolated and analysed as described below.

Preparation of the Formula (II) Using Human Hepatic Microsomes:

The incubation was performed with a final volume of 200 mL in a shakingwater bath at a temperature of approximately 37° C. Each incubationcontained approximately 400 ug/mL of microsomal protein (H51), 0.5 mL2mM the 4-hydroxyphenyl analog of pranlukast in a 16% w/v encapsinsolution containing 1.5% w/v sodium bicarbonate. The incubation volumewas adjusted to 150 mL with 50 mM potassium phosphate buffer (pH 7.4).Following a 5 minute pre-incubation at approximately 37° C., thereaction was initiated by the addition of 50 mL of pre-warmed cofactorsolution (approximately 1.7 mg NADP, 7.8 mg glucose 6-phosphate and 6units of glucose 6-phosphate dehydrogenase per mL of 2% (w/v) sodiumhydrogen carbonate). The reaction was terminated after 30 minutes byadding 200 mL of acetonitrile and vortex mixed. The precipitate waspelleted by centrifugation. The putative dihydroxymetabolite formed wassubsequently analysed and isolated as described below.

Isolation and Purification of the Formula (I) and Formula (II):

The in vitro samples (approximately 200 mL for Formula (I) andapproximately 400 mL for Formula (II) were freeze-dried overnight. Tothe dried solid, 15 mL of methanol was added and the solution wassonicated for approximately two minutes and then centrifuged (approx.8,000×g, 10 min) on a IEC Model K Centrifuge (International EquipmentCo., Needham Heights, Mass.). The supernatant was removed and theextraction repeated twice more. The supernatants were subsequentlycombined and the solvent was evaporated under a nitrogen stream at roomtemperature. The residue was reconstituted in 1.5 mL ofacetonitrile:water (2:1). The solution was aliquoted into 15 vials ofapprox. 100 uL solution each and was analyzed using the HPLC system andconditions described above. The eluent from repeat injections wascollected (from the UV detector outlet) into vials at 0.4 min intervalsusing a Foxy fraction collector (ISCO, Lincoln, Nebr.). The desiredfractions for collection were determined based on the UV chromatogram.The combined fractions were freeze-dried and the residue was loaded ontoa C18-Mega Bond Elut (Analytichem International, Harbor City, Calif.).The column was washed with water (approximately 4 mL) to remove residualsalt from the sample. The metabolite was then eluted from the columnwith methanol (approximately 4 mL). Solvent was removed from the sampleunder a stream of nitrogen and then lyophilized prior to storage for NMRanalysis. The purified sample was stored in a dessicator prior to NMRanalysis. This purification procedure was verified by using ¹⁴ C!SB205312 in a small scale incubation to ensure that all the extractionsteps were quantitative.

Qualitative HPLC-MS and HPLC-MS-MS Analysis

Sample Preparation:

Samples from the human S9 incubations were received as a 1:1 mixture ofincubation media and acetonitrile. In order to reduce the relativeorganic content to about 25%, the samples were diluted with an equalvolume of water prior to HPLC/MS and HPLC/MS/MS analysis. Samples fromthe microsomal incubations were also received as a 1:1 mixture ofincubation media and acetonitrile. The relative organic content washowever reduced to about 25% by evaporation following the centrifugationstep.

HPLC conditions for LC/MS analyses:

HPLC pumps: Hitachi L6200A and L6000

HPLC autosampler ThermoSeparations AS3000

Column Waters Symmetry C8 (150×3.9 mm), 5 um particle

size

Guard Column Waters Symmetry C8 (30×2.0 mm), 5 um particle

size

Solvent A: 10 mM ammonium formate pH 4.0

Solvent B: acetonitrile

HPLC flow rate: 1.0 mL.min⁻¹

Temperature: ambient

Split ratio into the MS: approximately 9.5:1

Gradient conditions:

    ______________________________________    Time (min)       % A    % B    ______________________________________    0                95     5    60               30     70    61               0      100    62               0      100    65               95     5    85               95     5    ______________________________________

Mass Spectrometry:

Mass Spectrometer: Finnigan TSQ700 Triple Quadrupole Mass Spectrometer

Data System: Personal DECStation running ICL (v.7.4) and ICIS(v.7.0)Ionization

Mode: Electrospray (operating in the negative ionization mode)

Scanning: 200-800 amu, 2 sec/scan (single quad scans)

Collision Gas: Argon, 2 mTorr

Collision Energy: 25 eV

As indicated above, the column effluent was split such that a flow of 50uL/min was directed into the mass spectrometer. The remainder wasdirected into the 125 uL solid flow cell of a Beckman 171 RadioisotopeDetector.

Multiple reaction monitoring was conducted for the human microsomalsamples. The following transitions, 496→292, for Formula (I) , and512→294, for Formula (II), were monitored by alternately allowing n/z496 and m/z 512 to pass through Q1 into the collision cell whilesimultaneously setting Q3 to allow only the appropriate fragment mass(292 or 294) to pass through to the detector.

Nuclear Magnetic Resonance (NMR) Spectroscopy

The proton NMR spectra of the pranlukast metabolites were measured at400.13 MHz, using a Bruker Instruments AMX400 spectrometer which wasequipped with an inverse gradient multinuclear probe maintained at 25°C. Samples ranging from 150 ug to 200 ug were dissolved in 0.5 mL ofdeuterated pyridine (Aldrich Chemical Co., Milwaukee, Wis.) usingtetramethylsilane as an internal reference. A spectral width of 5435 Hzwas measured using 32K data points. Each spectrum resulted from signalaveraging 48 scans. Proton nuclear Overhauser experiments (nOe) weremeasured by selectively irradiating the methine at 5.13 ppm and themethylene at 4.01ppm, along with an off-resonance control frequency,with low RF power for seven seconds prior to acquisition. For eachspectrum, 256 scans were signal averaged at the selected frequency, thenthe process repeated 8 times leading to 2048 scans per irradiation.Following Fourier transformation, difference spectra were generated bysubtracting the off-resonance control spectrum from each irradiatedspectrum.

In order to confirm proton connectivity, a COSY 2-dimensional experimentwas measured for the sample. The experiment from the AMX microprogramlibrary employed the "cosy" pulse program. The 512 spectra in the F2dimension were measured using 1024 data points over a spectral width of6024 Hz. The incrementable delay was increased in successive spectra toachieve a spectral width of 6024 Hz in the F1 dimension. For eachspectrum, 64 scans were signal averaged with a pulse delay of 1 sbetween successive spectra.

RESULTS Identification of Pranlukast Human Hepatic Metabolites

The negative electrospray ionization mass spectra of pranlukast andassociated compounds are characterized by a deprotonated molecule of thetype M-H!⁻. Collision induced dissociation of these compounds wasgenerally characterized by three predominant processes: loss of thetetrazole nitrogens as molecular N₂, loss of the butyl phenyl group, andamide bond cleavage. The means by which the butyl phenyl group was lostappeared to be sensitive to the degree, and possibly the position, ofhydroxylation. For example, following the loss of the tetrazole group byexpulsion of molecular nitrogen, pranlukast appeared to lose the butylphenyl group via homolytic bond cleavage (m/z 291). A homolytic processalso appeared to be responsible for the ion resulting from amide bondcleavage (m/z 171) for pranlukast. Incorporation of a single hydroxylgroup in either the 3- or 4-positions of the phenyl ring yielded astructure which favored heterolytic bond cleavage resulting in evenelectron ions. Thus, the loss of the butyl phenyl group and amide bondcleavage produced ions at m/z 292 and 172, respectively. Incorporationof a hydroxyl groups at both the 3- and 4-positions of the pheny ringalso yielded a structure in which the dominant fragmentation modesappeared to involve heterolytic bond cleavage. However, loss of thebutyl phenyl group in the case of the 3,4-dihydroxyphenyl analog ofpranlukast used as the standard, resulted in a ion at m/z 294. Thisfragmentation may be rationalized by the presence of the ortho-hydroxylgroups which could each transfer a hydrogen atom to the carbene(resulting from loss of the trazole nitrogens). Elimination of theresulting alkyl quinone would yield an ion at m/z 294.

Incubation of pranlukast with human liver S9 from donor H51 demonstratedthe formation of two major metabolites of pranlukast, nominated MM4 andMM5, which were both dependent on the presence of NADPH. Retention timesobserved in the ion chromatograms for m/z 498 and 482 (pranlukast)correlate well with those in the radiochromatogram. MM4 and MM5 showedparent ions at m/z 498 indicating that both metabolites were products ofmono-oxygenation. Note that the specific activity of the incubationsample was such that the predominant ionic species were observed at 2amu higher than the nominal ¹² C-monoisotopic mass. Therefore, in theCID spectra of the two monohydroxylated metabolites and the parentcompound, the precursor and fragment ions are all shifted by +2 amu dueto the predominance of the ¹⁴ C! label. The observed fragmentationpatterns are identical to the standards and confirmed the presence ofpranlukast, as well as two monohydroxylated metabolites.

The early eluting peak, MM4, demonstrated fragments of m/z 294 and 174,consistent with oxygen incorporation in the alkyl phenyl region of theparent molecule. The formation of a 3-hydroxyphenyl or4-hydroxyphenylcompound could be ruled out on the basis of retentiontime . The proton NMR spectrum of isolated MM4 was compared to theproton NMR spectrum of pranlukast. The aromatic regions were identicalexcept for slight differences in the chemical shifts of the protonsignals of the mono-substituted phenyl group. In the reference spectrum,the mono-substituted phenyl ring spin-system (H2"-H4") is contained inthe region from δ7.22 to δ7.38. The metabolite spectrum indicates thatthe multiplets for H4" and H2" range from δ7.30 to δ7.45, while H2" isshifted to δ7.69 relative to the reference spectrum consistent with achange at position 8'. The aliphatic region of the reference spectrum ofpranlukast displays H5' and H8' as triplets at δ3.9 1 and δ2.64,respectively. For the metabolite spectrum, the aliphatic region of thespectrum displays H8' as a one-proton doublet of doublets at δ5.12,which represents a dramatic down-field shift of ca. δ2.5. The chemicalshift of the oxymethylene, H5', is shifted about δ0.2. The data at thispoint indicate that hydroxylation occurred at position 8' in themetabolite. Further confirmation was obtained through decoupling and nOedifference experiments. In the decoupling experiments, both H8' and H5'were irradiated. Irradiation of H8', resulted in sharpening the doubletof multiplets at δ7.69, H2", indicating allylic coupling and providingadditional evidence that hydroxylation occurred at H8'. Irradiation ofH5' did not result in changes in the aromatic region of the spectrum.Irradiation at both positions, H8' and H5', resulted in changes in theregion of overlapping multiplets, δ2.2-δ2.0. For the nOe experiments,saturation of H5' enhanced H3' of the oxybenzamide group, whilesaturation of H8', resulted in the enhancement of H2" of themono-substituted phenyl group, confirming that the hydroxyl group islocated at position 8' Formula (I)!.

A novel dihydroxy metabolite was found in human fecal samples which didnot co-chromatograph with the 3,4-dihydroxyphenyl standard. Therefore,incubation of the 4-hydroxyphenyl pranlukast analog with human livermicrosomes and NADPH was conducted in order to confirm the identity ofthis metabolite. As illustrated by the reconstructed ion chromatogramsof m/z 496 (the 4-hydroxyphenyl analog) and m/z 512 (dihydroxylatedmetabolite), LC/MS analysis suggested that a metabolite withchromatographic retention time identitical to the novel fecal metabolitehad been generated in vitro. Background subtracted mass spectra derivedfrom the peaks in the ion chromatograms, were consistent with thepresence of the 4-hydroxyphenyl analog and a dihydroxylated metabolite.Further structural characterization of the metabolite was provided byMS/MS. The presence of m/z 292, as opposed to m/z 294, in the CIDspectrum of m/z 512 suggested a structure hydroxylated on the alkylchain as well as on the aromatic ring. Based on the metabolism ofpranlukast (vide supra), a likely site of hydroxylation of the4-hydroxyphenyl analog would be at the benzylic position. The proton NMRspectrum of MM9 was compared to the proton NMR reference spectrum of the4-hydroxyphenyl compound. The aromatic regions were identical except forthe proton signals of the 4-hydroxy-phenyl group. In the referencespectrum, the 4-hydroxy-phenyl ring, an A₂ B₂ spin-system including H2"and H3" protons, is contained in the region from δ7.25 to δ7.19. Theanalogous metabolite spectrum indicates a larger separation betweenprotons H2" and H3", which are located at δ7.60 and δ7.31, respectively.The relatively large shift for proton H2" (ca. δ0.36) relative to thesame signal in the reference spectrum is consistent with a change atposition 8'. The aliphatic region of the reference spectrum of the4-hydroxyphenyl compound displays H5' and H8' as triplets at δ3.91 andδ2.64, respectively. For the MM9 spectrum, however, the aliphatic regionof the spectrum displays H8' as a one-proton doublet of doublets atδ5.12, which represents a dramatic down-field shift of ca. δ2.5. Thechemical shift of the oxymethylene, H5', is shifted about δ0.2. Thechemical shift for H8' is consistent with the proton of a benzylicmethine bearing a hydroxyl group. Further confirmation of the positionof hydroxylation was obtained through decoupling and nOe differenceexperiments. In the decoupling experiments, both H8' and H5' wereirradiated. Irradiation of H8', resulted in sharpening the doublet atδ7.60, H2", indicating allylic coupling and providing evidence thatsupports hydroxylation at H8'. Irradiation of H5' did not result inchanges in the aromatic region of the spectrum. Decoupling at bothpositions, H8' and H5', resulted in changes in the region of overlappingmultiplets, δ2.2-δ2.0, which contains methylene protons H6' and H7'. Forthe nOe experiments, saturation of H5' enhanced H3' of the oxybenzamidephenyl ring, while saturation of H8', resulted in the enhancement of H2"of the 4-hydroxy phenyl group, confirming that the hydroxyl group islocated at position 8'.

SYNTHETIC EXAMPLES Example 1 Preparation of 4-Oxo-8-4-(4-Hydroxy-4-Phenylbutoxy)Benzoylamino!-2-Tetrazol-5-yl-4H-1-Benzopyran

2-Phenyl-2-(3-Chloropropyl)-1,3-Dioxolane

4-Chlorobutyrophenone (15.18 g, 0.083 moles), ethanediol (6.80 g, 0.110moles) and p-toluene sulphonic acid (0.20 g) were heated in toluene atreflux and water was removed from the reaction using a Dean and Starkapparatus. After 17 hours the solvent was removed by distillation invacuo to leave crude 2-phenyl-2-(3-chloropropyl)-1,3-dioxolane (21 g).

NMR (CDCl₃) ppm, 1.85, 2H, m; 2.03, 2H, m; 3.50, 2H, t; 3.74, 2H, m;3.98, 2H, m; 7.31, 3H, m; 7.42, 2H, dd.

Methyl 4-(4-oxo-4-Phenylbutoxy)Benzoate Ethylene Acetal.

Crude 2-phenyl-2-(3-chloropropyl)-1,3-dioxolane (21.0 g), potassiumcarbonate (14.0 g; 0.101 moles) and methyl 4-hydroxybenzoate (14.0 g,0.092 moles) were heated in dimethylformamide (100 ml) for 21 hours. Themixture was diluted with water (250 ml) and extracted withdichloromethane (500 ml). The dichloromethane extract was washed withsaturated sodium bicarbonate solution and then with water. The organicextract was concentrated in vacuo to give the product as a brown solid(27.3 g).

MS (e.i.) m/z (%) 77 (43), 105 (93), 149 (100), 265 (40), 342 (10).

NMR (CDCl₃) ppm, 1.89, 2H, m; 2.07, 2H, m; 3.79, 2H, t; 3.98, 3H, s;4.00, 4H, m; 6.86, 2H, d; 7.29-7.51, 5H, m; 7.95, 2H, d.

4-(4-Oxo-4-Phenylbutoxy)Benzoic Acid Ethylene Acetal.

Methyl 4-(4-oxo-4-phenylbutoxy)benzoate ethylene acetal (27.0 g),denatured alcohol (90 mls), water (90 ml) and sodium hydroxide (9.0 g,0.225 moles) were heated at reflux for 3 hours. The mixture was cooledto 20° C., diluted with water (250 ml) and extracted with toluene (2×250ml). The aqueous phase was acidified with concentrated hydrochloric acid(35 ml) and the precipitate filtered, washed with water and dried togive 4-(4-oxo-4-phenylbutoxy)benzoic acid ethylene acetal (21.1 g, 70%from 4-chlorobutyrophenone).

MS (Negative ion ionspray) m/z M-H!⁻ 327

NMR (DMSO-d₆) ppm, 1.72, 2H, m; 1.98, 2H, m; 3.70, 2H, t; 4.00, 4H, m;6.94, 2H, d; 7.23-7.96, 5H, m; 7.84, 2H, d; 12.5, 1H, br.s.

4-Oxo-8-4-(4-Oxo-4-Phenylbutoxy)Benzoylaminol-2-Tetrazol-5-yl-4H-1-BenzopyranEthylene Acetal.

4-(4-Oxo-4-phenylbutoxy)benzoic acid ethylene acetal (7.5 g, 0.023mole), thionyl chloride (3.26 g, 0.027 mole) and dimethylformamide (20mg) were heated in dichloromethane (75 mls) at reflux for 2 hours. Thesolvent was removed in vacuo and the residue, pyridine (5.38 g, 0.068mole) and 8-amino-4-oxo-2-tetrazol-5-yl-4H-1-benzopyran (SB-241906: 5.34g, 0.023 mole) were heated in toluene (50 mls) at 100° C. for 90minutes. The reaction was cooled to 20° C. and dilute hydrochloric acidadded. The crude product was filtered and the wet cake dissolved in warmmethanol (100 ml) and sodium acetate (2.0 g, 0.024 mole). Insolublematter was removed by filtration and the supernatant liquor acidifiedwith concentrated hydrochloric acid (2.5 mls). The precipitate wasfiltered, washed with methanol and dried to give the title compound(12.7 g 103%).

MS (Positive ion ionspray) m/z (%) 105 (18), 121 (12), 147 (100), 175(21), 191 (29), 496 (41), 512 (15), 540 (96).

NMR (DMSO-d₆) ppm, 1.77, 2H, m; 2.02, 2H, m; 3.70, 2H, t; 7.07, 2H, d;7.13, 1H, s; 7.30-7.47, 5H, m; 7.56, 1H, t; 7.91, 1H, d; 8.03, 2H, d;8.29, 1H, d; 7.96, 1H, s.

4-Oxo-8-4-(4-Oxo-4-Phenylbutoxy)Benzoylamino!-2-Tetrazol-5-yl-4H-1-Benzopyran

4-Oxo-8-4-(4-oxo-4-phenylbutoxy)benzoylamino!-2-tetrazol-5-yl-4H-1-benzopyran₋₋ethylene acetal (12.5 g, 0.023 moles) water (20 ml) and p-toluenesulphonic acid (200 mgs) were heated in acetone (700 ml) at reflux for16 hours. The reaction was cooled to 20° C. and filtered to give4-oxo-8-4-(4-oxo-4-phenylbutoxy)benzoylamino!-2-tetrazol-5-yl-4H-1-benzopyran(5.04 g, 44%).

MS (Positive ion ionspray) m/z (%) 105 (22), 147 (100), 175 (12), 266(6), 294 (18), 350 (5), 468 (6), 496 (46).

NMR (DMSO-d₆) ppm, 2.13, 2H, m; 3.23, 2H, t; 4.18, 2H, t; 7.13, 3H, m;7.50-7.69,4H, m; 7.91,2H, d; 8.02, 4H, m; 8.29, 2H, d; 9.98, 1H, s.

4-Oxo-8-4-(4-Hydroxy-4-Phenylbutoxy)benzoylamino!-2-Tetrazol-5-yl-4H-1-Benzopyran

Sodium borohydride (1.45 g) was added, in portions to 4-oxo-8-4-(4-oxo-4-phenylbutoxy)benzoylamino!-2-tetrazol-5-yl-4H-1-benzopyran(5.0 g, 0.010 mole) in methanol (100 ml) at 20° C. The reaction wasdiluted with water (400 ml) and 1 M hydrochloric acid (35 ml) added. Thesolid was filtered, washed with water and dried at 80° C. to give4-oxo-8-4-(4-hydroxy-4-phenylbutoxy)benzoylamino!-2-tetrazol-5-yl-4-benzopyran(4.95 g, 98%).

Mpt. 198-205° C.

IR (Nujol mull) cm⁻¹, 3300 (m), 1659 (s), 1642 (s), 1605 (m), 1584 (m),1527 (m), 1506 (s), 1463 (s), 1430 (s), 1377 (s), 1289 (s), 1252 (s),1179 (m), 1036 (m), 765 (m).

MS (Positive ion ionspray) m/z (%) 131 (100), 147 (26), 175 (28), 228(7), 251 (31), 396 (6), 424 (100), 435 (19), 452 (15), 480 (50), 498(28).

NMR (Pyridine-d₅) ppm 2.00-2.20, 4H, m; 4.03, 2H, m; 5.09, 1H, m; 7.06,2H, d; 7.29-7.47, 4H, m; 7.70, 2H, m; 8.17, 1H, d; 8.51, 2H, d; 8.94,1H, d; 10.25, 1H, s.

What is claimed is:
 1. A compound of formula I in essentially pure form##STR2## or a pharmaceutically acceptable salt thereof; or an enantiomerthereof, a hydrate, solvate or a polymorph thereof.
 2. A compound ofFormula II in essentially pure form ##STR3## or a pharmaceuticallyacceptable salt thereof; or an enantiomer, a solvate, a hydrate or apolymorph thereof.
 3. A pharmaceutical composition comprising a compoundof Formula I in essentially pure form ##STR4## or a pharmaceuticallyacceptable salt thereof; or an enantiomer, a solvate, a hydrate or apolymorph thereof; and a pharmaceutically acceptable excipient.
 4. Apharmaceutical composition comprising a compound of Formula II inessentially pure form ##STR5## or a pharmaceutically acceptable saltthereof; or an enantiomer, a solvate, a hydrate or a polymorph thereof;and a pharmaceutically acceptable excipient.
 5. A method for treatingasthma comprising administering to a mammal in need thereof apharmaceutically acceptable composition comprising a pharmaceuticallyacceptable excipient and an essentially pure form of formula (I) or(II). ##STR6##
 6. A method for manufacturing a pharmaceuticalcomposition comprising mixing a compound according to claims 1 or 2 witha pharmaceutically acceptable excipient.